TS-071 is a novel, potent and selective renal sodium-glucose cotransporter 2 (SGLT2) inhibitor with anti-hyperglycaemic activity.

BACKGROUND AND PURPOSE: The renal sodium-glucose cotransporter 2 (SGLT2) plays an important role in the reuptake of filtered glucose in the proximal tubule and therefore may be an attractive target for the treatment of diabetes mellitus. This study characterizes the pharmacological profile of TS-071 ((1S)-1,5-anhydro-1-[5-(4-ethoxybenzyl)-2-methoxy-4-methylphenyl]-1-thio-D-glucitol hydrate), a novel SGLT2 inhibitor in vitro and in vivo. EXPERIMENTAL APPROACH: Inhibition of glucose uptake by TS-071 was studied in CHO-K1 cells stably expressing either human SGLT1 or SGLT2. Single oral dosing studies were performed in rats, mice and dogs to assess the abilities of TS-071 to increase urinary glucose excretion and to lower plasma glucose levels. KEY RESULTS: TS-071 inhibited SGLT2 activity in a concentration-dependent manner and was a potent and highly selective inhibitor of SGLT2. Orally administered TS-071 increased urinary glucose excretion in Zucker fatty rats and beagle dogs at doses of 0.3 and 0.03 mg·kg(-1) respectively. TS-071 improved glucose tolerance in Zucker fatty rats without stimulating insulin secretion and reduced hyperglycaemia in streptozotocin (STZ)-induced diabetic rats and db/db mice at a dose of 0.3 mg·kg(-1). CONCLUSION AND IMPLICATIONS: These data indicate that TS-071 is a potent and selective SGLT2 inhibitor that improves glucose levels in rodent models of type 1 and 2 diabetes and may be useful for the treatment for diabetes mellitus.

Prevention of hypercholesterolemia and atherosclerosis in the hyperlipidemia- and atherosclerosis-prone Japanese (LAP) quail by taurine supplementation.

The effects of taurine supplementation on the serum cholesterol levels and the progression of atherosclerosis were investigated in the hyperlipidemia- and atherosclerosis-prone Japanese (LAP) quail. The ingestion of a high-cholesterol diet containing 1% cholesterol by LAP quails for 60 days resulted in a marked elevation in serum non-HDL cholesterol and triglyceride, as well as severe aortic lesions with lipid droplets. An immunohistochemical study showed that the lesion consisted of mainly lipid-rich macrophages and T cells. Sixty-day taurine supplementation (1% in drinking tap water) to LAP quails fed high-cholesterol diet containing 1% cholesterol significantly reduced serum non-HDL cholesterol from 4,549 to 2,350 mg/dl. The serum triglyceride level also decreased after taurine supplementation from 703 to 392 mg/dl. Although the HDL cholesterol level significantly decreased due to the high-cholesterol diet, it recovered to the control level fed a regular diet in response to taurine. Bile acid production was stimulated and hepatic cholesterol was reduced by taurine supplementation. A quantitative analysis using aortic cross-sections showed that areas of oil-red O positive lipid accumulation significantly decreased by 74% after taurine supplementation. These results demonstrated the lipid-lowering and anti-atherosclerotic effects of taurine in a diet-induced hyperlipidemic LAP quail model. The prevention of atherosclerosis by taurine is mainly attributed to an improvement in the serum cholesterol and triglyceride levels, which may be related to changes in the hepatic cholesterol metabolism.

Pathological role of angiostatin in heart failure: an endogenous inhibitor of mesenchymal stem-cell activation.

OBJECTIVE: Recently, a clinical trial was initiated to evaluate the efficacy of transendocardial transplantation of autologous bone marrow-derived mesenchymal stem cells (MSC) for the treatment of heart failure (HF). Because some HF patient-derived sera did not induce proliferation of autologous MSC, the present study aimed to elucidate humoral factors in sera that attenuate MSC activation and to investigate the role of these humoral factors in the pathogenesis of HF. METHODS AND RESULTS: Inhibitory effects present in serum were analysed by culturing human MSC with sera from 10 HF patients (FS <25%, BNP >100 pg/ml) and four healthy control subjects. Among the patients, two sera from HF patients showed significant inhibitory activity on MSC proliferation. Protein array and ELISA analysis revealed that these sera contained high levels of angiostatin as well as the active form of matrix metalloproteinase (MMP)-9, which generates angiostatin. Angiostatin significantly inhibited the proliferation and migration of cultured human MSC and increased their apoptosis in a dose-dependent manner. In a rat HF model, serum levels of angiostatin and MMPs increased, but treatment with an MMP inhibitor suppressed these increases. CONCLUSIONS: The results suggest that angiostatin, which can attenuate the activity of MSC, might play a role in the progression of HF.

Plasma protein S activity correlates with protein S genotype but is not sensitive to identify K196E mutant carriers.

BACKGROUND: Protein S (PS) is an anticoagulant protein that functions as a cofactor for activated protein C (APC), and congenital PS deficiency is a well-known risk factor for the development of deep vein thrombosis (DVT). Recently, we and others identified the K196E missense mutation in the second epidermal growth factor-like domain of PS as a genetic risk factor for DVT in the Japanese population. The incidence of this mutation is high in the Japanese population. OBJECTIVES: In the present study, we investigated the relationship between plasma PS activity and the presence of the K196E mutation. PATIENTS AND METHODS: We measured PS activity as a cofactor activity for APC in 1,862 Japanese individuals and determined the PS K196E genotype in this population. RESULTS: Individuals heterozygous for the mutant E-allele had lower plasma PS activity than wildtype subjects (mean +/- SD, 71.9 +/- 17.6%, n = 34 vs. 87.9 +/- 19.8%, n = 1,828, P < 0.0001). However, the PS activity of several heterozygous individuals (n = 8) was greater than the population average. In contrast, multiple wildtype subjects (n = 26) had PS activity less than 2 SD below the population mean, indicating that other genetic or environmental factors affect PS activity. CONCLUSIONS: Plasma PS activity itself is not suitable for identifying PS 196E carriers and other methods are required for carrier detection.

Genetic variations and haplotype structures of the ABCB1 gene in a Japanese population: an expanded haplotype block covering the distal promoter region, and associated ethnic differences.

As functional ABCB1 haplotypes were recently reported in the promoter region of the gene, we resequenced the ABCB1 distal promoter region, along with other regions (the enhancer and proximal promoter regions, and all 28 exons), in a total of 533 Japanese subjects. Linkage disequilibrium (LD) analysis based on 92 genetic variations revealed 4 LD blocks with the same make up as previously described (Blocks -1, 1, 2 and 3), except that Block 1 was expanded to include the distal promoter region, and that a new linkage between polymorphisms -1,789G>A in the distal promoter region and IVS5 + 123A>G in intron 5 was identified. We re-assigned Block 1 haplotypes, and added novel haplotypes to the other 3 blocks. The reported promoter haplotypes were further classified into several types according to tagging variations within Block 1 coding or intronic regions. Our current data reconfirm the haplotype profiles of the other three blocks, add more detailed information on functionally-important haplotypes in Block 1 and 2 in the Japanese population, and identified differences in haplotype profiles between ethnic groups. Our updated analysis of ABCB1 haplotype blocks will assist pharmacogenetic and disease-association studies carried out using Asian subjects.

Haplotype structures of the UGT1A gene complex in a Japanese population.

Genetic polymorphisms of UDP-glucuronosyltransferases (UGTs) are involved in individual and ethnic differences in drug metabolism. To reveal co-occurrence of the UGT1A polymorphisms, we first analyzed haplotype structures of the entire UGT1A gene complex using the polymorphisms from 196 Japanese subjects. Based on strong linkage disequilibrium between UGT1A8 and 1A10, among 1A9, 1A7, and 1A6, and between 1A3 and 1A1, the complex was divided into five blocks, Block 8/10, Block 9/6, Block 4, Block 3/1, and Block C, and the haplotypes for each block were subsequently determined/inferred. Second, using pyrosequencing or direct sequencing, additional 105 subjects were genotyped for 41 functionally tagged polymorphisms. The data from 301 subjects confirmed the robustness of block partitioning, but several linkages among the haplotypes with functional changes were found across the blocks. Thus, important haplotypes and their linkages were identified among the UGT1A gene blocks (and segments), which should be considered in pharmacogenetic studies.

Activation of distinct signal transduction pathways in hypertrophied hearts by pressure and volume overload.

OBJECTIVE: Two types of hemodynamic overload, pressure and volume overload, result in morphologically distinct types of cardiac remodeling. We explored the possibility that distinct hemodynamic overload may differentially activate the signal transduction pathway. METHODS: Pressure and volume overload were induced by thoracic aortic banding and carotid-jugular shunt formation in rabbits, respectively. Phosphorylation activities of mitogen-activated protein (MAP) kinase families, Akt, and signal transducer and activator of transcription (STAT) 3 in the left ventricular myocardium were determined by Western blotting using phospho-specific antibodies and were compared between hypertrophied hearts by pressure and volume overload. RESULTS: Pressure and volume overload produced concentric and eccentric cardiac hypertrophy in rabbits, respectively. In pressure-overloaded hearts, extracellular signal-regulated kinase (ERK) 1/2, p38 MAP kinase, and STAT3 were transiently activated prior to hypertrophic changes. In contrast, activation of ERK1/2, but not p38 MAP kinase and STAT3, was observed only at 12 weeks after shunt surgery. Pressure overload evoked short and biphasic activation of Akt at 15 min and 1 day after aortic banding. In contrast, volume overload induced sustained activation of Akt from 1 day to 1 week. Concordant phosphorylation of downstream targets of Akt, glycogen synthase kinase-3beta (GSK-3beta) and p70 ribosomal S6 kinase (p70(S6K)), in response to Akt activation was observed at 15 min after pressure overload. However in volume-overloaded hearts, phosphorylation of GSK-3beta and p70(S6K) was observed at 6 weeks and at 6 and 12 weeks, respectively, and was not coincident with Akt activation. These findings suggest that phosphorylation of GSK-3beta and p70(S6K) is regulated by an alternative pathway other than Akt in volume-overloaded hearts. CONCLUSION: Pressure and volume overload-induced cardiac hypertrophy is associated with distinct patterns of activation of signal transduction pathways. These data may suggest that stimulus-specific heterogeneity in the signaling pathway plays a role in determining the type of cardiac hypertrophy.

Fatty acid metabolism assessed by 125I-iodophenyl 9-methylpentadecanoic acid (9MPA) and expression of fatty acid utilization enzymes in volume-overloaded hearts.

BACKGROUND: The peroxisome proliferator-activated receptor (PPAR) alpha is a member of the nuclear receptor superfamily and regulates gene expression of fatty acid utilization enzymes. In cardiac hypertrophy and heart failure by pressure-overload, myocardial energy utilization reverts to the fetal pattern, and metabolic substrate switches from fatty acid to glucose. However, myocardial metabolism in volume-overloaded hearts has not been rigorously studied. The aim of the present study was to examine fatty acid metabolism and protein expressions of PPARalpha and fatty acid oxidation enzymes in volume-overloaded rabbit hearts. METHODS: Volume-overload was induced by carotid-jugular shunt formation. Sham-operated rabbits were used as control. Chronic volume-overload increased left ventricular weight and ventricular cavity size, and relative wall thickness was decreased, indicating eccentric cardiac hypertrophy. (125)I-iodophenyl 9-methylpentadecanoic acid (9MPA) was intravenously administered, and animals were sacrificed at 5 min after injection. The 9MPA was rapidly metabolized to iodophenyl-3-methylnonanoic acid (3MNA) by beta-oxidation. Lipid extraction from the myocardium was performed by the Folch method, and radioactivity distribution of metabolites was assayed by thin-layer chromatography. The protein was extracted from the left ventricular myocardium, and levels of PPARalpha and fatty acid oxidation enzymes were examined by Western blotting. RESULTS: Myocardial distribution of 9MPA tended to be more heterogeneous in shunt than in sham rabbits (P = 0.06). In volume-overloaded hearts by shunt, the conversion from 9MPA to 3MNA by beta-oxidation was faster than the sham-control hearts (P < 0.05). However, protein levels of PPARalpha and fatty acid utilization enzymes were unchanged in shunt rabbits compared with sham rabbits. CONCLUSIONS: These data suggest that myocardial fatty acid metabolism is enhanced in eccentric cardiac hypertrophy by volume-overload without changes in protein expressions of PPARalpha and fatty acid utilization enzymes. Our data may provide a novel insight into the subcellular mechanisms for the pathological process of cardiac remodelling in response to mechanical stimuli.

Population-based distribution of plasminogen activity and estimated prevalence and relevance to thrombotic diseases of plasminogen deficiency in the Japanese: the Suita Study.

Reduced plasminogen activity with a normal level of antigen is commonly observed in Japanese individuals. The first reported patient with plasminogen deficiency was accompanied with deep vein thrombosis. The present study examines whether heterozygous or homozygous deficiency of plasminogen is a risk factor for thrombotic disease. This study measures the plasminogen activity of 4517 individuals in the general population, determines the cut-off to define plasminogen deficiency, and identifies plasminogen deficiencies in the control groups and thrombotic disease groups. In another study, we examined the phenotypes of consecutive patients with homozygous plasminogen deficiency detected in our hospital. We found 173 and two of 4517 individuals to have heterozygous and homozygous deficiency with normal plasminogen antigen level, respectively, and 19 to have heterozygous deficiency with reduced antigen levels. The incidence of plasminogen deficiency in an age- and sex-matched control group (13/324, 4.01% for deep vein thrombosis or 13/330, 3.94% for stroke) selected from the 4517 individuals was not significantly different from those in patients with deep vein thrombosis (3/108, 2.78%) or cardioembolic stroke (6/110, 5.55%). Among 19 patients with homozygous plasminogen deficiency showing about 10% plasminogen activity, none had deep vein thrombosis. These findings indicate that neither heterozygous nor homozygous plasminogen deficiency constitutes a significant risk factor for thrombotic disease.

Left ventricular diastolic dysfunction in patients with bronchial asthma with long-term oral beta2-adrenoceptor agonists.

BACKGROUND: Long-term administration of oral beta(2)-adrenergic receptor agonists (beta(2)-AG) in patients with bronchial asthma (BA) causes disastrous events such as sudden death and heart attack. However, long-term effects of beta(2)-AG on cardiac function have not been previously quantified. METHODS: Seventy-four patients with BA with regular long-term use of oral beta(2)-AG (group A) and 69 patients with BA without beta(2)- AG (group B) were examined in medical records of outpatient clinics from 1985 to 1999. In the prospective study, echocardiography was performed in 48 consecutive patients from January to April 1999. There were 26 patients with regular oral use of beta(2)-AG (group a) and 22 patients without beta(2)-AG (group b). Twenty-one age-matched normal volunteers without heart or pulmonary diseases were used as control subjects (group c). Oral beta(2)-AG was withdrawn from remedies in 11 patients of group a, and echocardiographic studies were repeated 2 weeks after its cessation. RESULTS: Events related to heart failure were more frequently seen in group A than in group B (14% vs 1%, P <.01). The echocardiographic study showed that indexes of left ventricular diastolic but not systolic function were significantly deteriorated in group a, along with a markedly reduced level of plasma norepinephrine concentration (P <.05 vs groups b and c). When heart rate was adjusted to 90 beats/min during isoproterenol infusion, left ventricular diastolic function remained deteriorated in group a (P <.05 vs groups b and c). In 11 patients of group a, the cessation of beta(2)-AG for 2 weeks resulted in an improvement of left ventricular diastolic function and in an increase of plasma norepinephrine level (P <.01). CONCLUSIONS: Long-term use of oral beta(2)-AG impaired left ventricular diastolic function in patients with BA, and the cessation of beta(2)-AG reversed diastolic pump performance to the normal level.

Reverse redistribution of 99m Tc-sestamibi after direct percutaneous transluminal coronary angioplasty in acute myocardial infarction: relationship with wall motion and functional response to dobutamine stimulation.

Reverse redistribution (RR) of 99mTc-sestamibi is observed after direct percutaneous transluminal coronary angioplasty (PTCA) in acute myocardial infarction (AMI). The purpose of this study was to clarify the functional characteristics of myocardial segments with RR after direct PTCA in AMI. Thirty patients with AMI who had undergone direct PTCA were examined. Myocardial perfusion tomography with 99mTc-sestamibi and low dose dobutamine echocardiography were performed within 2 weeks of the onset. The 99mTc-sestamibi images were obtained 1 and 3 h after tracer administration. The left ventricle was divided into nine segments, and regional 99mTc-sestamibi uptake and clearance were quantitatively evaluated in each segment. RR was defined as a decrease in 99mTc-sestamibi uptake of >10% on 3 h delayed images compared with the 1 h early images. The left ventricle in the echocardiographic images was also divided into nine segments corresponding to the scintigraphic images, and regional wall motion was assessed in the resting condition as the baseline and during dobutamine administration (5-10 microg x kg(-1) x min(-1)). Out of a total of 270 myocardial segments, 111 segments were perfused by the culprit coronary artery and were defined as ischaemic segments. There were 25 segments with RR and 86 segments without RR in the ischaemic myocardium. Enhanced clearance of 99mTc-sestamibi was observed in ischaemic segments with RR (P<0.001). Echocardiography demonstrated that 24 out of 25 segments with RR and 61 out of 86 segments without RR had wall motion abnormalities. Dobutamine infusion improved wall motion in 20 (83%) of the 24 dysfunctional segments with RR and 33 (54%) of the 61 dysfunctional segments without RR (P<0.02). These findings suggest that RR indicates reversible functional abnormalities associated with preserved contractile reserve in response to dobutamine. The early and delayed imaging of 99mTc-sestamibi provides useful information regarding the residual viability of the dysfunctional myocardium in AMI patients.

Role of calcineurin in insulin-like growth factor-1-induced hypertrophy of cultured adult rat ventricular myocytes.

The present study examined the role of calcineurin in insulin-like growth factor (IGF)-1-induced hypertrophy in primary cultures of adult rat ventricular myocytes (ARVM), prepared from the ventricles of 14-16-week-old male Sprague-Dawley rats. The effects of several humoral factors, including phenylephrine, angiotensin II, endothelin-1, IGF-1 and interleukin-6, on the morphology of ARVM were studied. Myocyte surface area was significantly increased by IGF-1 (2,268 +/- 571 to 3,018 +/- 836 microm2, p < 0.01), but not by other humoral factors. This hypertrophic effect of IGF-1 was blocked by genistein (tyrosine kinase inhibitor), PD98059 (MEK inhibitor). These findings suggest that IGF-1 produces ARVM hypertrophy by a tyrosine kinase-MEK mediated pathway as has been reported in neonatal cardiomyocytes. IGF-1-mediated ARVM hypertrophy was also attenuated by cyclosporine A (calcineurin inhibitor), and staurosporine and chelerythrine (protein kinase C inhibitors). IGF-1 markedly increased calcineurin activity (8.7 +/- 1.2 to 98.0 +/- 54.3 pmol x h(-1) mg(-1), p < 0.01), and this activation was completely blocked by pre-treatment with cyclosporine A (8.5 +/- 11.4pmol x h(-1) x mg(-1), p < 0.01) and chelerythrine (2.3 +/- 2.7 pmol x h(-1) mg(-1), p < 0.01). It appears that IGF-1 activates calcineurin by a protein kinase C-dependent pathway. Increased mRNA expression of atrial natriuretic factor by IGF-1 was inhibited by cyclosporine A (p < 0.01). The findings indicate that IGF-1 induces ARVM hypertrophy by protein kinase C and calcineurin-related mechanisms. The fact that elevated calcineurin activity and induced atrial natriuretic factor mRNA expression by IGF-1 were blocked by cyclosporine A further supports the hypothesis that calcineurin is critically involved in IGF-1-induced ARVM hypertrophy.

Long-term prognosis after recovery from myocardial infarction: a community-based survey in Yamagata, Japan.

OBJECTIVE: To assess the long-term prognosis after recovery from acute myocardial infarction (AMI) in the general population in Japan. PATIENTS AND METHODS: Among the 575,000 inhabitants of the Yamagata metropolitan area, a total of 117 patients suffered from first their AMI from April to December 1993. Thirteen patients (11%) died within four weeks after the onset. Of the remaining 104 patients, 101 (mean age, 69+/-12 years) were followed for an average of 65+/-5 months. RESULTS: Twenty-seven of the 101 patients (27%) died during the follow-up period. Compared with survivors, the patients who died were significantly older at the onset of AMI (74+/-12 vs. 67+/-12 years, p<0.01). More diabetic patients than non-diabetic patients died (42 vs. 21%, p<0.05) because of the higher frequency of non-cardiac deaths (29 vs. 11%, p<0.05). The total number of deaths of cardiac origin, including sudden deaths, was 11 (40%) and was lower than the number of definite non-cardiac deaths (n=15). The time from the onset of AMI to death was significantly shorter in cases of cardiac death than in cases of non-cardiac death (median, 16 vs. 45 months, p<0.01). Among non-cardiac deaths, deaths due to lung cancer and cerebral infarction were notable in men (standardized mortality ratio 278) and women (571), respectively. CONCLUSION: Non-cardiac death during long-term follow-up after AMI was more frequent than death of cardiac origin. Thus, preventive measures, including early treatment of complicating diseases, must be implemented to improve the long-term prognosis of patients with myocardial infarction.

Impaired distensibility of the left ventricle after stiffening of the right ventricle.

Acute and chronic alterations of right ventricular (RV) wall properties can change left ventricular (LV) performance. We investigated whether and how stiffening of the RV free wall alters LV diastolic distensibility. We used cross-circulated isolated hearts, in which the LV and RV were independently controllable. Stiffness of the RV free wall was altered by intramuscular injections of glutaraldehyde into the RV free wall after right coronary artery ligation. We measured circumferential and longitudinal regional lengths in the septum and LV free wall. During data acquisition, RV volume was held constant. After the RV free wall was stiffened by glutaraldehyde, the LV diastolic pressure-volume relation shifted upward and became steeper. Importantly, stiffening of the RV free wall increased the diastolic regional area in the septum and LV free wall under constant LV volume. The augmented regional dimensions may result in enhanced regional tension under constant LV volume and may be related to the observed increase in LV diastolic intracavitary pressure. The impaired LV diastolic distensibility by stiffening of the RV free wall may be at least partly explained by myocardial stretch, probably due to LV deformation.

A novel pathophysiologic phenomenon in cachexic patients with chronic obstructive pulmonary disease: the relationship between the circadian rhythm of circulating leptin and the very low-frequency component of heart rate variability.

Cachexic patients with chronic obstructive pulmonary disease (COPD) show abnormalities of the autonomic nervous system (ANS), neuroendocrine function, and energy expenditure. Leptin has been implicated in the regulation of ANS, neuroendocine function, and thermogenesis in humans. We assessed the physiologic significance of the circadian rhythm of circulating leptin using power spectrum analysis of heart rate variability (HRV) in nine cachexic male patients with COPD, eight noncachexic patients with COPD, and seven healthy control subjects. A diurnal pattern of 24-h leptin levels was present in both the control subjects (analysis of variance [ANOVA]; F = 7.80, p < 0.0001) and noncachexic COPD patients (F = 9.29, p < 0.0001), but was strikingly absent in the cachexic COPD patients (F = 2.09, p = NS). Analysis of HRV demonstrated that the diurnal rhythm of 24-h very low frequency (VLF; 0.003 to 0.04 Hz) showed significantly identical fluctuations with those of 24-h leptin levels, in all of the three groups (r = 0.388, p < 0.0001). Because VLF has been considered to reflect neuroendocrine and thermoregulatory influences, these data may suggest that the loss of circadian rhythm of circulating leptin has clinical importance in the pathophysiologic features in cachexic patients with COPD.

Alterations in the inotropic responses to forskolin and Ca2+ and reduced gene expressions of Ca2+-signaling proteins induced by chronic volume overload in rabbits.

Volume overload results in eccentric cardiac hypertrophy, but it is still unknown how this mechanical overload modulates the inotropic response to exogenous Ca2+ or adenylyl cyclase stimulation. Inotropic responsiveness in vivo and the levels of gene expression of Ca2+ signaling proteins were studied in rabbit hearts hypertrophied as a result of volume overload at 4 and 12 weeks after arteriovenous shunt formation. In sham-operated control rabbits, left ventricular (LV)+dP/dt was augmented in response to graded doses of CaCl2. Dose-related changes of LV+dP/dt to CaCl2 were attenuated significantly in shunt rabbits with volume overload. Forskolin dose-dependently augmented LV+dP/dt in sham rabbits, which was also attenuated significantly in rabbits with volume overload. The mRNA levels of dihydropyridine receptor, Na+/Ca2+ exchanger, sarcoplasmic reticulum Ca2+-ATPase, and ryanodine receptor decreased significantly at 4 and 12 weeks in the volume-overload rabbits compared with the sham rabbits, but the mRNA levels of phospholamban and calsequestrin remained unchanged. Chronic volume overload alters contractile responsiveness to Ca2+ or adenylyl cyclase stimulation, and downregulation of steady state mRNA levels of Ca2+ signaling proteins might be, at least in part, related to this pathologic process.

Dobutamine stress echocardiography for the diagnosis of myocardial viability: assessment of left ventricular systolic velocities in longitudinal axis by pulsed Doppler tissue imaging.

Dobutamine (DOB) stress two-dimensional echocardiography is an established method for the detection of viable myocardium, but conventional assessment of wall motion is subjective. We measured quantitatively the left ventricular systolic velocities along the longitudinal axis by pulsed Doppler tissue imaging (DTI). In 30 patients with previous myocardial infarction, pulsed DTI focused on the infarct area was performed from an apical two- or four-chamber view before and during DOB (10 microg/kg/min) stress one day before coronary angioplasty. We calculated peak systolic velocity (S), regional pre-ejection period (PEP, the time interval from the onset of QRS to the onset of systolic wave) and regional ejection time (ET). Left ventriculography was obtained before and 3 months after coronary angioplasty to assess regional wall motion. Improvement of abnormal wall motion was observed in 19 patients (group P) but not in 11 (group N). Group P had significantly larger S and smaller PEP/ET than group N during DOB stress, although there were no significant differences in these indices between the groups at baseline. As a consequence, group P had a significantly larger percent change in S and a smaller percent change in PEP/ET than group N (164+/-39 vs 117+/-20% and 88+/-17 vs 116+/-29%, respectively, p < 0.01). It is suggested that the quantitative measurement of longitudinal systolic velocities during DOB stress by DTI is useful for the precise assessment of myocardial viability.

Percutaneous in vivo gene transfer to the peripheral lungs using plasmid-liposome complexes.

The purpose of this study was to investigate a new method of in vivo gene transfer to the lung parenchyma by the percutaneous approach. The plasmid that contains the gene for firefly luciferase driven by a cytomegalovirus (CMV) promoter (pCMVL) in combination with cationic lipids was percutaneously injected into the lung parenchyma. Luciferase activities were localized to the lobes of the lung where the plasmids with cationic lipids were injected. Percutaneous injection of the plasmid containing the human endothelin-1 (hET-1) gene driven by a CMV promoter (pRc/CMVhET-1) in combination with cationic lipids into the lungs caused pulmonary fibrosis localized to the injection site in the peripheral lungs. We concluded that percutaneous in vivo gene transfer to the lungs is a unique and important approach to introduce exogenous gene expression in the limited area of the lung parenchyma. This method of gene transfer will be applicable for human gene therapy for targeted areas of peripheral lung and will also be useful to assess the function of the proteins expressed by a gene in the local area of the lungs.